There is a growing body of evidence suggesting that some Chlamydia proteins are able to gain access to the host cell cytosol to promote survival of these obligate intracellular organisms. I propose to identify Chlamydia proteins that are translocated from the vacuole where Chlamydia reside into the host cell cytosol or the vacuolar membrane. This will be done by generating a C. trachomatis library where individual Chlamydia proteins are fused to known peptide epitopes recognized by CD8+ T cells. Translocation of these fusion proteins can then be monitored by recognition of the epitope tags by epitope-specific CD8+ T cells. Proteins identified using this method will be further characterized to define the role of each protein in directing an immune response against Chlamydia and in establishing this organism's intracellular niche. The experimental system proposed here is a unique approach for identifying potential virulence determinants in this elusive pathogen. Identification of Chlamydia proteins that are translocated into the host cell cytosol will yield insight into C. trachomatis pathogenesis and cell biology. In addition, translocated proteins that are targets of the immune system can be considered as components of experimental Chlamydia vaccines.